1. Immunology and Inflammation
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Protective function and durability of mouse lymph node-resident memory CD8+ T cells

  1. Scott M Anthony
  2. Natalija Van Braeckel-Budimir
  3. Steven J Moioffer
  4. Stephanie van de Wall
  5. Qiang Shan
  6. Rahul Vijay
  7. Ramakrishna Sompallae
  8. Stacey M Hartwig
  9. Isaac J Jensen
  10. Steven M Varga
  11. Noah S Butler
  12. Hai-Hui Xue
  13. Vladimir P Badovinac  Is a corresponding author
  14. John T Harty  Is a corresponding author
  1. Department of Pathology, The University of Iowa, United States
  2. Department of Microbiology and Immunology, The University of Iowa, United States
  3. Center for Discovery and Innovation, Hackensack Meridian Health, United States
  4. Interdisciplinary Graduate Program in Immunology, The University of Iowa, United States
Research Article
Cite this article as: eLife 2021;10:e68662 doi: 10.7554/eLife.68662
7 figures, 3 tables and 1 additional file

Figures

CD103+ memory CD8+ T cells are generated in draining lymph nodes (LNs) during localized but not systemic infections.

(A) C57BL/6 mice were infected IN with X31 (H3N2); mice were sacrificed 90 days post-infection; non-draining cervical lymph nodes (cLN) or lung-draining mediastinal lymph nodes (mLNs) were harvested and analyzed by flow cytometry. (B) Representative plots of % of CD69 and CD103 expression (left) in NP366 tetramer+ IV- memory CD8+ T cells from the cLN or mLN and cumulative data (right). n = 3–5 mice/group. Representative of three independent experiments. Bars denote mean values, dots represent independent mice. ****p<0.0001, Students t-test. (C) Mice were seeded with 104 naive P14 cells and infected IN with either PR8-GP33 (H1N1) or Vac-GP33. 30 days post-infection, draining mLNs were isolated and CD69+ CD103+ P14 Trm populations were evaluated (D). Representative plots (left), cumulative data (right). Representative of two independent experiments, n = 5 mice/group. Error bars represent mean ± SD. ****p<0.0001, Students t-test. (E) Mice were seeded with 104 naive P14 cells and infected IP with LCMV Armstrong. 30 days post-infection LNs (mLN and iLN) were isolated and evaluated for the frequency (F) of CD69+/CD103+ P14 s.

Lung-draining LN Trm cells mediate local protective immunity.

Mice were seeded with 104 naive or 105 3M P14 cells and IN infected with PR8-GP33 virus. At 50 days post-infection, frequency of 1M or 4M P14s were measured in the peripheral blood (A). At 100 days post-infection, mLNs were harvested and the numbers of total (B) and CD103+ CD69+ (C) P14s were determined. Representative of three independent experiments, n = 4–5 mice/group. Error bars represent mean ± SD, *p<0.05, **p<0.01, Students t-test in (A–C). LCMV challenge (D,E). Naïve, 1M, or 4M mice were infected with LCMV-Armstrong (2.0 × 105 PFU/mouse i.p.); 72 hr post LCMV challenge, mLN (D) and SP (E) were harvested and individually evaluated for LCMV titers by plaque assay. FTY720 treatment impact on LCMV challenge (F). Naïve, 1M, or 4M mice were infected with LCMV-Armstrong (2.0 × 105 PFU/mouse i.p.) and treated with vehicle or FTY720 daily for 72 hr. 72 hr post LCMV challenge, mLN (F) were harvested and individually evaluated for LCMV titers by plaque assay. Dotted line denotes limit of detection. One representative of 2–3 independent experiments is shown, n = 3–5 mice/group. Error bars represent mean ± SEM (G) or mean ± SD (H). NS = not significant, *p<0.05, one-way ANOVA.

Figure 3 with 1 supplement
Repeated antigen stimulation extends the survival of LN Trms.

Mice were seeded with 104 naive or 105 3M P14 cells and IN infected with PR8-GP33 virus. At indicated time points, mLN (A) was harvested and total numbers of 1M (white) and 4M (black) P14 cells were evaluated. Representative of three independent experiments, n = 4 mice/group/time point. Error bars represent mean ± SD. ***p<0.001, ****p<0.0001, two-way ANOVA with Sidak’s multiple comparison test. Representative plots (B) and cumulative results (C) of 1M and 4M CD69+ CD103+ P14 Trm cells in mLN evaluated at indicated time points. Representative of three independent experiments, n = 4 mice/group/time point. Error bars represent mean ± SD. *p<0.05, **p<0.01, ****p<0.0001, two-way ANOVA with Sidak’s multiple comparison test.

Figure 3—figure supplement 1
CD103+ LN Trm cells are not present within non-draining iLN.

Mice were seeded with 104naive P14 or 105 3M P14 cells and IN infected with PR8-GP33 virus. At indicated time points, iLN (A) were harvested and total numbers of 1M (white) and 4M (black) P14 cells were evaluated. Representative of three independent experiments, n = 4 mice/group/time point. Error bars represent mean ± SD. ***p<0.001, ****p<0.0001, two-way ANOVA with Sidak’s multiple comparison test. Representative plots of 1M and 4M CD69+ CD103+ P14 Trm cells in iLN (B) evaluated 30 and 150 days post-infection. Representative of three independent experiments, n = 4 mice/group/time point.

Figure 4 with 2 supplements
Repeated antigen stimulation increases granzyme production of LN Trms.

Mice were seeded with 104 naive or 105 3M P14 cells and IN infected with PR8-GP33 virus. At >60 days post-infection, mLNs were harvested, stimulated with cognate (GP33) peptide in the presence of BFA for 5 hr, and ICS was performed to evaluate the frequency of GrzA+ and GrzB+ fractions of IV CD103 or CD103+ 1M or 4M by flow cytometry. Representative flow plots (A), cumulative frequencies (B), and total numbers per mLN (C) are shown. Representative of two independent experiments, n = 4–5 mice/group. Error bars represent mean ± SD. *p<0.05, one-way ANOVA in (B), *p<0.05, Students t-test in (C).

Figure 4—figure supplement 1
Repeated influenza stimulation reduces cytokine production, but does not affect degranulation capacity of LN Trm cells.

Mice were seeded with 104naive P14 or 105 3M P14 cells and IN infected with PR8-GP33 virus. At 60 days post-infection, mLN and spleen were harvested and cells analyzed for peptide-stimulated cytokine production and degranulation. (A) Gating strategy for analysis of IV P14s. Single-cell suspensions were stimulated with cognate peptide (GP33) in the presence of BFA for 5 hr and ICS was performed to evaluate the frequency of CD107+, IFNγ+, TNF+, and IL-2+ fractions of IV CD103 or CD103+ 1M or 4M by flow cytometry. Representative flow plots of IFNg, TNF, and IL-2 (B), cumulative bar graphs denoting frequencies of CD107+ (C, left) and CD107+ IFNg+ (C, right) are displayed. Representative of two independent experiments, n = 4–5 mice/group. NS = not significant, *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001; two-way ANOVA with Sidak’s multiple comparison test.

Figure 4—figure supplement 2
Repeated antigen stimulation alters the phenotype of LN Trm cells.

Mice were seeded with 104naive P14 or 105 3M P14 cells and IN infected with PR8-GP33 virus. At >90 days post-infection, iLN (top) and mLN (bottom) were harvested and the frequency of 1M and 4M P14 cells evaluated. (A) Representative data examining CD62L expression; cumulative frequency data (middle); cumulative total number data (right). Representative of three independent experiments, n = 4 mice/group. Error bars represent mean ± SD. ***p<0.001, ****p<0.0001 one-way ANOVA. At >75 days post-infection, mLN were harvested from 1M and 4M mice and the relative expressions of CD122 (B), Eomes (C), CD49a (D), CXCR6 (E), and CXCR3 (F) by CD103+ (red) or CD103 (blue) P14s were evaluated. Representative histograms (top) and cumulative data (bottom) are displayed. Representative of three independent experiments, n = 4–5 mice/group. Error bars represent mean ± SD. NS = not significant, *p<0.05, ****p<0.0001, one-way ANOVA.

Repeated antigen stimulation alters localization of LN Trm cells.

(A–D) Mice containing mixed congenically distinct populations of 1M (CD90.1/.1, seeded with 104 naive P14) and 4M (CD90.1/.2 seeded with 105 3M P14) P14s were injected with bolus IV administration of CD90.1-PE (red) and CD90.2-APC (white). Approximately 5 hr post-injection, organs were isolated and two-photon microscopy was performed on whole mLN (A–B) or iLN (C–D) explants ex vivo. The LN surrounding collagen capsule (pseudocolored blue) was captured with secondary harmonic generation (SHG). Representative of two independent experiments, n = 3–4 mice/group.

Residential nature of 1M and 4M LN Trm cells primed by influenza infection.

(A) 90 days after IN PR8-GP33 infection, mice bearing Thy1.1/1.1 1M P14 (green; 1M mice seeded with 104 naive P14) cells were joined by parabiotic surgery with mice bearing Thy1.1/1.2 4M P14 (purple, 4M mice seeded with 105 3M P14). Three weeks later parabionts were analyzed. (B) Abundance of 1M (green) and 4M (purple) P14 cells in iLNs of 1M (top row) and 4M (bottom row) parabiotic mice. Representative plots (left), cumulative data (right). Representative of two independent experiments, n = 4 parabionts/experiment. Error bars represent mean ± SD. ****p<0.0001, t-test. (C) Abundance and distribution of 1M (green) and 4M (purple) Trm P14 cells expressed as a % of the total Trm population (CD69+/CD103+) in mLN of 1M (top row) and 4M (bottom row) parabiotic mice. Representative plots (left), cumulative data (right). Representative of two independent experiments, n = 4 parabionts/experiment. Error bars represent mean ± SD. Two-way ANOVA with Sidak’s multiple comparison test. Q1(1M) vs Q1(4M) ****p<0.0001; Q2(1M) vs Q2(4M) ****p<0.0001; Q1(1M) vs Q1(4M) ****p<0.0001.

Figure 7 with 1 supplement
Splenic 4M cells express a core Trm signature.

Mice were seeded with 104 naive P14 or 105 3M P14 cells and IN infected with PR8-GP33 virus. At 22–30 days post-infection, IV exclusion was performed and negatively enriched pooled groups of spleens (3–5 spleens/sample, n = 3) or mLNs (15–25 mLNs/sample, n = 2) were stained for CD8α, CD90.1, CD69, and CD103. Bulk RNAseq was performed on RNA from sort-purified spleen samples (20k IV, CD69/CD103 cells/sample) or mLN Trm samples (2–5k IV CD69+/CD103+ cells/sample). (A) Heatmaps of 1300 most differentially expressed genes (log2FC > 1.5, p<0.05) between 1M and 4M LN Trms are plotted from the four respective groups of samples. The six core signature sets of genes offset to the right were derived from unbiased hierarchical clustering. (B) Volcano plot of 4061 differentially expressed genes between 1M and 4M splenic memory P14 cells (log2FC > 1.5, p<0.05). (C) GSEA of core Trm genes defined in Table 2 from splenic 1M and 4M populations separated into respective upregulated (top) and downregulated (bottom) gene sets in regard to annotated expression in Trms. (D) Heatmap of a core set of selected Trm genes (as in C) within 1M and 4M splenic populations.

Figure 7—figure supplement 1
Gating strategy of FACS for RNAseq.

Mice were seeded with 104 naive P14 or 105 3M P14 cells and IN infected with PR8-GP33 virus. At 23–30 days post-infection, mLN and spleen were pooled, harvested, negatively enriched, and sorted for bulk RNAseq. (A) Gating strategy for analysis of 1M or 4M splenic IV Tcirc P14s. (B) Gating strategy for analysis of 1M or 4M IV mLN Trm P14s.

Tables

Table 1
Samples for RNAseq, cell number, and RNA quality.
RNAseq sampleInformation
Sample info(pg/µL) RNA concentrationSample volume (µl)Total ng, RNA per tube(RIN) RNA
Integrity Number as determined by Agilent
RNA ratio
1M Sp DN #16181710.506N/A2
1M SP DN #26781711.526N/A2
1M SP DN #310181717.306N/A2
4M Sp DN #16531711.1019.32.2
4M SP DN #26921711.7649.62.3
4M SP DN #310281717.4769.42.1
1M mLN Trms78171.3269.21.8
1M mLN Trms139233.1977.41.8
4M mLN Trms12001720.49.11.8
4M mLN Trms8181713.9068.71.8
Table 2
RNAseq heatmaps and GSEA full data.

RNAseq expression related to Figure 6.

GeneSp1M-1Sp1M-2Sp1M-3Sp4M-1Sp4M-2Sp4M-3
Neurl3−1.448372−0.7443671.4137260.0291060.5586490.191258
Ctla4−1.472581−0.893646−0.0856310.7646740.937260.749924
Fn1−0.4761−0.7844470.313751−0.47611.898997−0.4761
Arl4c−0.219927−0.637316−0.954088−0.3682041.8395740.33996
Pdcd1−0.974857−1.5210790.2718570.6475150.790750.785815
Bcl2−1.381925−0.6532171.527735−0.0806610.0199930.568075
Rgs2−1.527485−0.9837760.3442620.8180420.64780.701157
Rgs16−1.15537−1.037752−0.4692270.8871620.8360290.939158
Fcgr2b−0.959014−0.670621−1.0688590.8536310.7526441.092219
Cd244−0.4632−0.776816−1.377560.8844620.8769980.856116
Slamf7−1.139613−1.165538−0.2949170.9038170.8606080.835643
Atf3−0.779708−0.779708−0.7797080.5172440.1206881.701193
Il2ra−1.322889−1.1275480.0218610.7336150.8892190.805742
Zeb2−1.316736−1.060834−0.1249860.8439570.6924070.966192
Nr4a2−1.166233−1.064171−0.3910730.701880.8612061.058391
Itga60.103598−1.3548661.208944−1.0450110.4770860.610248
Chn1−0.408248−0.408248−0.4082482.041241−0.408248−0.408248
Itga4−1.533817−0.9350030.9560910.5902660.2548590.667605
Cd44−0.687106−0.641904−1.1524410.1386321.0616341.281186
Sema6d−1.095775−1.095775−0.2772191.3041330.3151540.849482
Dusp2−1.284804−1.2482050.3633330.5169720.7117240.94098
Bcl2l1−1.599476−0.9207340.6505490.5632170.6603480.646096
Fgf13−1.184461−1.3528960.333820.5806070.8363260.786603
Cxcr3−1.558265−0.9347180.893920.3683230.6701380.560601
Fabp5−0.937223−0.580515−1.1615530.9662390.9879020.72515
Tnfsf10−0.757581−1.3349−0.5291280.7828340.7450861.093689
Rorc−0.833666−0.8336660.249621−0.8336660.7267271.52465
Cd101−1.953973−0.1224770.3286940.4243730.6217660.701617
Csf1−1.089549−0.89601−0.7350680.9050820.8727740.94277
S1pr1−1.414071−1.105490.2617920.6953590.8344680.727942
Usp33−1.470917−1.0454780.7252380.8698960.3258550.595405
Bach2−1.176816−0.990541.5231350.4171380.352504−0.125421
Aqp3−1.330326−1.2165940.8337990.3714890.7627890.578843
Coro2a−1.303957−1.114023−0.0624950.7849560.8721980.823321
Tnfsf8−1.655898−0.0802021.4780250.0135870.258145−0.013656
Dmrta1−0.845576−1.1498191.698186−0.0559750.0941920.258992
Jun−1.376227−1.1624850.3131260.6818020.7650770.778706
Zfp683−1.593545−0.9021610.4625860.7343250.4911770.807619
Runx3−1.150841−1.3271610.9872230.1855120.489350.815917
Tnfrsf1b−1.135037−1.4268120.6221790.4988930.6889430.751834
Abcb1a−1.375583−1.001921−0.1094720.6917160.8660810.929179
Cd36−1.132023−1.1320230.6470121.3507240.414674−0.148364
Fosl20.574051−1.743753−0.6171940.2605880.8530120.673296
Cd38−1.726006−0.5467150.1692240.3422290.7683550.992914
Klf3−1.580977−0.9485170.6123430.6342950.6230610.659795
Cxcl9−0.779708−0.7797080.5172441.7011930.120688−0.779708
Tgfbr3−1.496104−1.0031560.2722290.6115540.8420890.773388
Dtx1−1.323911−0.5323751.651323−0.2711030.0796770.39639
Vps37b−1.327024−1.2482280.7315730.5238880.6731870.646604
Hsph1−0.89509−1.269058−0.4820190.7634310.9215050.961231
Chn2−1.469041−1.0398230.3667150.5657770.9665510.609821
Il12rb2−1.512436−0.863636−0.0441590.7171790.8419540.861097
Cd8b1−0.897526−1.057029−0.743550.6505281.072770.974808
Cd8a−1.369955−0.899035−0.2931710.6914080.9589220.91183
Sema4f−0.711384−1.581909−0.1307660.7076070.8411390.875313
Mxd1−1.636166−0.8353040.8202570.4200780.5204510.710683
Bhlhe40−1.377693−0.960477−0.1943660.7949750.8766390.860922
Klrg1−0.649531−0.92849−1.1279760.8804010.9458220.879774
Lag3−1.314079−1.2506610.476270.5621760.7368420.789452
Klre1−0.817609−0.924946−0.9914910.9008790.8931070.940061
Klrd1−0.82806−1.237129−0.6056830.7790210.844551.0473
Klrc2−1.060961−0.22419−1.1578391.3128170.3187380.811434
Klrc1−0.456844−0.628525−1.481540.7993080.8740630.893538
Dusp16−1.549192−0.590124−0.3043610.4825390.9402861.020853
Emp1−0.820283−1.020728−0.886020.7900460.9617560.975229
Fosb−1.192929−1.192929−0.1047190.8179430.6426771.029956
Nkg70.8566620.552759−1.936070.0028260.4555850.068237
Ppp1r15a−1.743276−0.6907680.6518690.5833820.597130.601662
Swap70−1.185293−1.2736270.028840.7604660.8368270.832787
Il4ra−1.240991−1.2702771.0219410.408520.3806980.70011
Il21r−1.376141−1.1768480.8582310.4300210.6305420.634194
Itgal−1.41421−1.1391990.448350.5758170.778120.751121
Itgax−1.299963−1.141223−0.0059960.7301090.8436860.873388
Bag3−1.261842−1.3025850.5799480.4716190.8192370.693622
Adam8−1.448605−0.760408−0.3550880.8022970.9089790.852826
Ifitm2−1.9315680.697426−0.2219550.3543910.5721630.529542
Ifitm1−0.116103−0.604934−1.6390240.6020240.99390.764137
Ifitm3−0.399411−1.471071−0.006047−0.1649961.5523870.489138
Ifitm10−1.619476−0.8103670.1984430.6304750.8339450.76698
Ifngr1−1.689527−0.7242360.5428520.317250.8735660.680095
Prdm1−0.763724−0.527892−1.2666071.198390.9740680.385764
Prf1−1.450717−1.0221220.1298330.7324090.8193840.791213
Gzmm−0.148129−1.7688640.896369−0.2978850.8686970.449812
Gadd45b−1.732358−0.682270.4054530.5920530.6236960.793427
Phlda1−1.336075−1.1975980.2864570.7830690.8143780.649768
Ifng−1.270643−0.812118−0.5874440.8720870.9420330.856084
A430078g2−0.768864−1.6724110.3529950.7627010.5466240.778955
Rasa3−1.298438−1.2682760.4562230.6121020.7407350.757654
Dusp4−0.013625−1.524651−0.248373−0.0136251.6047240.19555
Lpl−0.751462−0.751462−0.751462−0.0584560.5961171.716724
Klf2−0.890686−1.058738−0.6228390.3040691.262581.005613
Inpp4b−1.411831−1.1229560.9179490.5487020.4084870.65965
Dnajb1−1.351654−1.1870330.503080.4560150.9562110.623381
Junb−1.248952−1.3241030.753570.5085820.6790450.631859
Cdh1−1.503072−0.8545781.2390260.4980730.178620.441931
Itgb1−1.070625−1.246026−0.2450610.6698110.9083230.98358
Fut11−1.566705−0.9073430.2644370.6404650.7106430.858503
Gzmc−1.129587−1.1295871.3937450.7015020.0819630.081963
Gzmb−1.019828−0.852547−0.8603090.8606050.9368480.935231
Tmem123−0.486668−0.568492−1.3832660.2892731.3328770.816276
Icam1−1.395117−1.1776560.6226490.5928310.7357180.621575
S1pr5−1.401126−1.0706170.1072560.687380.8265630.850543
Gm10080−0.187622−0.228053−1.7977390.5849550.7341870.894272
Smad3−1.296473−1.2540740.4365540.4947560.8176830.801554
Anxa2−1.161681−1.045596−0.4440540.7777620.9357550.937813
Cx3cr1−1.267574−1.170718−0.0230680.7782250.8460350.837099
Ccr8−1.030745−1.030745−0.6493170.8594990.9020930.949215
Crr9−1.163685−1.1636850.5181730.0593331.3692190.380645
Cxcr6−1.008609−0.713182−0.9986560.855040.9865420.878867
Ccr1−0.959366−0.959366−0.044992−0.4601371.1489551.274906
Ccr2−1.244353−0.427864−0.8351070.6475980.4702741.389451
Ccr5−1.267395−1.1977810.0848130.6002190.8304190.949725
Adam19−1.693324−0.3535171.3074940.4079040.361716−0.030273
Havcr2−0.267637−1.661471−0.4792170.7470140.8198720.841438
Itgae0.265873−0.131851−1.8906340.2878541.0309810.437777
Traf4−1.26168−1.3170190.6640030.6564930.7005340.557669
Ccl50.7178050.069708−1.9657560.1554630.6964470.326333
Ccl9−0.096406−1.204224−1.1769340.6856581.0956860.69622
Ccl30.3217410.441288−2.0338210.3472050.5573510.366236
Ccl4−0.900786−1.071795−0.7437440.7708110.9760860.969428
Wfikkn2−1.521007−0.4642331.525421−0.0379380.1519380.345819
Tbx21−1.188999−1.375880.4753450.6103450.7590990.720091
Arl5c−1.071685−1.4812440.6221130.5211950.7559160.653704
Stat3−1.33085−1.2448270.5684210.5758720.7221030.709281
Icam2−1.427743−1.0742860.3079950.5011990.9659030.726932
Cmah−1.619268−0.8200811.0124380.4813390.4439190.501653
Fam65b−1.435265−1.0876510.2783840.7565490.736230.751753
Irf4−0.942848−1.402093−0.1664210.7376520.8741450.899565
Ly86−1.176696−1.1766960.2714780.7022550.0930941.286564
Nfil3−1.352986−1.2106740.4426960.6649490.7807730.675242
Cdc14b−1.660352−0.625526−0.0189910.6824790.6244630.997927
Naip3−0.645497−0.645497−0.645497−0.6454971.2909941.290994
Elovl7−0.792302−0.420558−1.3505781.1072280.9343820.521828
Gzma−0.686737−0.814558−1.1971820.8487370.9410360.908703
Gzmk−1.406303−0.870165−0.2784020.7648090.911270.878791
Itga1−1.285811−1.102208−0.1132170.8349490.7035020.962785
Rhob−0.230141−1.8690040.1305540.3631980.712530.892863
Id2−1.046589−0.909419−0.7604470.7602240.9149861.041244
Adam40.587047−1.9859930.480645−0.0352460.5870470.3665
Fos−0.879435−1.290058−0.4740260.8821880.816450.94488
Il7r−0.170366−0.087313−1.8259590.4237860.640091.019763
Ly6c1−0.753065−1.398044−0.2146370.1747511.0678361.123158
Ly6c2−1.276681−1.2094690.159690.5481360.8961530.882171
Il2rb−1.273008−1.302080.7056620.5058920.6653740.698162
Bin2−1.441078−1.1226640.5463090.6004360.7095770.70742
Nr4a1−1.212606−1.3122521.0272330.5543660.4620630.481197
Itga5−0.995646−0.9956461.538635−0.3969880.6811750.16847
Litaf−1.67839−0.6921260.1531890.8174330.8273320.572561
Klhl6−1.527286−0.9083520.1379480.5615230.8883020.847865
Bcl6−1.042258−1.315998−0.1400060.8510541.0252310.621977
Tigit−0.636695−0.492403−1.4529250.801080.9728090.808135
Sidt1−1.416356−0.9179451.3523630.3209310.3085180.352489
Ccr6−0.408248−0.408248−0.408248−0.4082482.041241−0.408248
Hspa1a−1.489338−1.057890.5465750.592070.6008620.807722
Tnf−1.932303−0.2261970.372220.5082530.6663420.611686
Tnfsf9−0.721049−1.122053−0.8693070.8717380.995940.84473
Qpct−0.9723320.746934−0.972332−0.7738320.9191291.052433
Slc3a2−1.412175−1.036270.0688120.6417120.9199150.818006
Dtx4−1.8958210.0310870.9854320.2063820.6235260.049393
Dusp5−1.204462−1.231643−0.0095550.6557360.907920.882003
Aff30.3341580.9341361.302955−0.695519−1.131655−0.744075
Icos1.4711370.8413930.145304−0.792242−1.047218−0.618376
Ikzf2−0.1656711.7049780.658251−0.793835−0.609887−0.793835
Cxcr40.8691150.9193330.906884−0.886976−1.182769−0.625587
Cd550.6944670.4440261.397141−0.459842−1.074231−1.001562
Rgs10.282992−0.0288891.3206630.669655−1.444527−0.799896
Sell0.3430050.5773991.57097−0.879322−0.741027−0.871025
Xcl11.1580690.7267650.823087−0.933921−0.948108−0.825893
Slamf60.8781150.6480261.158166−0.969274−0.688009−1.027024
Dapl10.958760.9531710.822988−0.894936−0.945047−0.894936
Mal0.278549−1.1663880.89838−0.0247421.180589−1.166388
Pmepa11.0009431.530105−0.429028−0.700673−0.700673−0.700673
Fabp4−0.4082482.041241−0.408248−0.408248−0.408248−0.408248
Skil1.478111.07101−0.815616−0.553424−0.577221−0.602859
Il20.8430971.080960.786459−0.913213−0.735725−1.061578
Foxo11.3980041.047696−0.261312−1.167157−0.508615−0.508615
Il6ra0.1924770.1232631.813008−0.551343−0.551343−1.026062
Npr1−0.4082482.041241−0.408248−0.408248−0.408248−0.408248
Lef1−0.577413−0.1225822.008562−0.513026−0.534901−0.26064
Lpar3−0.600795−0.6007951.779753−0.600795−0.6007950.623426
Ifi44−1.5221881.3705390.77727−0.414406−0.162093−0.049122
Nr4a31.658887−1.4347770.298688−0.213395−0.096008−0.213395
Tlr11.087014−1.7597010.693949−0.332077−0.046720.357535
Cxcl100.819849−1.5536171.2170930.268993−0.179362−0.572955
Cd27−0.8127020.7360121.686726−0.76644−0.35287−0.490726
Cd91.1398530.9630040.584072−0.83021−0.83021−1.02651
Cd690.6246430.930421.125309−0.853084−0.712926−1.114363
Isg201.3934780.9346870.197602−0.930029−0.737091−0.858648
Rgs100.966341.0376460.595869−0.585759−0.644411−1.369686
Themis−0.6592091.957135−0.786652−0.207908−0.035484−0.267882
Fyn1.0665841.3465470.054683−0.726098−0.837588−0.904128
Egr21.036188−1.6368371.073379−0.057847−0.260015−0.154868
Timp3−0.4082482.041241−0.408248−0.408248−0.408248−0.408248
Dusp6−0.2059910.0851391.646718−1.49606−0.014903−0.014903
Crtam1.0792031.145370.408387−1.043006−0.745838−0.844116
Cxcr5−1.4752450.9949791.2045650.116051−0.50998−0.330371
Eomes−0.0051120.3622011.517082−1.5599440.048375−0.362601
Tcf7−0.5629370.0666071.970539−0.522208−0.289205−0.662796
Ccr7−0.893954−0.1633251.969748−0.395389−0.21246−0.30462
Cd861.3808651.183911−0.546108−0.52407−0.777066−0.717531
Btla0.9717060.8747180.860138−0.76399−1.171717−0.770855
Sik10.1951640.1951640.1951641.1168090.195164−1.897467
Epcam1.0638440.4984880.93633−1.165034−1.165034−0.168594
Egr1−0.013398−0.6993011.896141−0.6993010.151141−0.635282
Ifit31.1146240.96780.615644−0.896586−0.812924−0.988558
Key resources table
Reagent type (species) or resourceDesignationSource or referenceIdentifiersAdditional
information
Strain, strain background (Mus musculus)C57BL6/JJackson LaboratoryStock No: 000664
(RRID:IMSR_JAX:000664)
Strain, strain background (Mus musculus)B6.PL(84NS)/CyJackson LaboratoryStock No: 000983
(RRID:IMSR_JAX:000406)
C57BL6/J
Thy1.1
Strain, strain background (Mus musculus)B6.Cg-Tcratm1MomTg(TcrLCMV)327Sdz
(P14)
Jackson LaboratoryStock No: 37394-JAX
(RRID:IMSR_TAC:4138)
Strain, strain background (Mus musculus)Thy1.1/1.1- B6.Cg-Tcratm1Mom Tg(TcrLCMV)327SdzThis paperThy1.1/1.1 P14Can be acquired through lab contact or breeding of above commercially available strains
Strain, strain background (Mus musculus)Thy1.1/1.2- B6.Cg-Tcratm1Mom Tg(TcrLCMV)327SdzThis paperThy1.1/1.2 P14Can be acquired through lab contact or breeding of above commercially available strains
Strain, strain background (Influenza A virus)Recombinant influenza A/PR/8/34 expressing (H1N1) GP33-41Laidlaw et al. Cooperativity Between CD8+ T Cells, Non- Neutralizing Antibodies, and Alveolar Macrophages Is Important for Heterosubtypic Influenza Virus Immunity. Plos Pathog. 9(3) e1003207 (2013).PR8-GP33Can be acquired through lab contact.
Strain, strain background (Influenza A virus)Recombinant influenza A/X-31 (H3N2) expressing GP33-41Laidlaw et al. Cooperativity Between CD8+ T Cells, Non- Neutralizing Antibodies, and Alveolar Macrophages Is Important for Heterosubtypic Influenza Virus Immunity. Plos Pathog. 9(3) e1003207 (2013).X31-GP33Can be acquired through lab contact.
Peptide, recombinant proteinGP33-44AnaSpecCatalog #:
AS-61296
AntibodyCD11a
(rat monoclonal)
BiolegendM17/4
(AB_312776)
FACs
(1:100)
AntibodyIFNγ
(rat monoclonal)
eBioscienceXMG1.2
(AB_465410)
FACs
(1:100)
AntibodyCD8a
(rat monoclonal)
eBioscience53–6.7
(AB_1853141)
FACs
(1:100)
AntibodyThy1.1
(mouse monoclonal)
eBioscienceOX-7
(AB_2201314)
FACs
(1:100)
AntibodyThy1.2
(rat monoclonal)
eBioscience30-H12
(AB_1853152)
FACs
(1:100)
AntibodyCD45.2
(mouse monoclonal)
eBioscience104
(AB_469724)
FACs
(1:100)
AntibodyCD103
(hamster monoclonal)
Biolegend2E7
(AB_469040)
FACs
(1:100)
AntibodyCD69
(hamster monoclonal)
BiolegendH1.2F3
(AB_1853105)
FACs
(1:100)
AntibodyCD44
(rat monoclonal)
Biolegend1M7
(AB_223593)
FACs
(1:100)
AntibodyCD62L
(rat monoclonal)
BiolegendMEL-14
(AB_1853103)
FACs
(1:100)
AntibodyKLRG1
(mouse monoclonal)
eBioscience2F1
(AB_540279)
FACs
(1:100)
AntibodyCX3CR1
(mouse monoclonal)
eBioscienceSA011F11
(AB_2565701)
FACs
(1:100)
AntibodyCXCR3
(Armenian hamster monoclonal)
eBioscienceCXCR3-173
(AB_1210593)
FACs
(1:100)
AntibodyEomesodermin
(rat monoclonal)
eBioscienceDan11mag
(AB_11042577)
FACs
(1:100)
AntibodyTNF
(rat monoclonal)
eBioscienceMP6-XT22
(AB_465416)
FACs
(1:100)
AntibodyIL-2
(rat monoclonal)
BiolegendJES6-5H4
(AB_315298)
FACs
(1:100)
AntibodyGranzyme A
(mMouse monoclonal)
Biolegend3G8.5
(AB_2565308)
FACs
(1:100)
AntibodyGranzyme B
(rat monoclonal)
Biolegend12F9B65
(AB_2564373)
FACs
(1:100)
AntibodyBrdU
(mouse monoclonal)
BiolegendBu20a
(AB_1595472)
FACs
(1:100)
Commercial assay or kitFoxp3/Transcription Factor Staining Buffer SetInvitrogen00-5523-00
Software, algorithmGraphPad PrismGraphPad Prism 8Version 8.4.2 (464)
(RRID:SCR_002798)

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